Download DNA Test for PRA in the MLHD

Primary Lens Luxation
Cathryn Mellersh
Animal Health Trust
22nd November, 2009
Collaboration & Acknowledgements
•
David Sargan (University of Cambridge)
•
David Gould (Davies Veterinary Specialists)
•
AHT Ophthalmologists & BVA Eye Panelists
•
Owners and breeders who have donated DNA samples from their dogs
•
Kennel Club Charitable Trust, American Kennel Club Canine Health
Foudation, Breed Clubs & individuals.
Talk Layout
•
Brief overview of Primary Lens Luxation.
•
DNA – what it is, what it does, and how mutations in DNA cause inherited
disease.
•
How a mutation progresses through a pedigree.
•
How a mutation is tracked down
•
PLL mutation and DNA test
– what the results mean
– mutation frequency in the population
– general advice
•
Vitreal leakage
Primary Lens Luxation
Zonules
In the normal eye the lens is
held in place by numerous
fibres, called zonules.
Lens luxation occurs when
all of these fibres are broken
and the lens becomes loose
within the eye, where it can
move forwards or (less
commonly) backwards in the
eye.
PLL
Anterior lens luxation
Enucleated eye, due
to anterior lens
luxation and
secondary glaucoma.
PLL
• Primary Lens Luxation is a well-recognised, painful & potentially
blinding condition known to affect many terrier and terrier-type
breeds.
• Believed to be inherited as an autosomal recessive condition in
all breeds studied.
• Difficult to eliminate from at risk breeds because:
– Clinically unaffected carriers can pass mutation on to offspring
– Condition is mid-late onset, so affected dogs often bred with
prior to onset of clinical signs
DNA test required
DNA
•
DNA is a very long molecule & is found in virtually every cell of the body.
•
The canine genome consists of approximately 2,500,000,000 nucleotides of DNA
•
If each nucleotide was 1mm long the canine genome would stretch from Land’s
End to John O’Groats and back.
•
ALL the DNA is copied every time a cell divides
•
DNA is responsible for every aspect of you and your dog that is not controlled by
the environment.
Coding DNA
DNA
C
G
A
T
G
T
C
A
A
T
G
C
A
C
C
G
C
T
A
C
A
G
T
T
A
C
G
T
G
G
C
G
U
G
G
Transcription
mRNA
G
C
U
A
C
A
G
U
U
A
Translation
Protein
A
T
V
T
W
DNA
•
DNA is a code/blueprint for every physical characteristic of a dog that is not
determined by the environment.
•
The code is determined by the order of nucleotides along the DNA.
•
ALL 2,500,000,000 nucleotides are copied every time a cell divides.
•
Mistakes that arise are called MUTATIONS.
•
Most mutations that arise are repaired whereas a tiny minority persist.
•
Most mutations have no effect, whereas some can be advantageous.
•
Some mutations have a deleterious effect & cause inherited disease.
*
Chromosomes
Sperm
Egg
fertilization
X
38 autosomes & 1 X chromosome
Y
38 autosomes & 1 X or Y chromosome
38 pairs of autosomes
& 1 pair sex
chromosomes
Spontaneous Mutation
X
Recombination
X
Propagation
X
Inheritance of Mutation Down a Pedigree
Parents
G-Parents
G G-Parents
G G G-Parents
Shared Region of DNA
Affected
Mutation Identification
Affected
Unaffected
PLL research
•
Research to identify mutation responsible for PLL initiated ~ 10 years ago.
•
DNA from affected & unaffected dogs was analysed to identify region of the genome
that was homozygous in all affected dogs, and different in the unaffected dogs.
•
– MBTS
40 cases & 11 controls
– LHs
22 cases & 18 controls
– X breeds
5 cases
…to cut a very long story short we identified the mutation responsible for PLL in
Miniature Bull Terriers & Lancashire Heelers in summer 2009.
•
*
It is a single nucleotide substitution.
Breeds carrying PLL mutation
•
Miniature Bull Terrier
•
Jagd Terrier
•
Lancashire Heeler
•
Volpino Italiano
•
Jack Russell Terrier
•
Australian Cattle Dog
•
Parson Russell Terrier
•
Yorkshire Terrier
•
Patterdale Terrier
•
Sealyham Terrier
•
Chinese Crested Dog
•
Tibetan Terrier
Between Breeds
DNA Test Results
DNA test developed and made available October 19th, 2009.
Clear
Carrier
Genetically Affected
PLL DNA Test
• http://www.aht.org.uk/genetics_pll.html
• Kits for taking cheek swabs are available by phoning
+44 (0)1638 555621 or via e-mail to [email protected].
• Further information [email protected]
PLL DNA Test Results
• CLEAR: these dogs have two normal copies of DNA. Our research
has demonstrated clear dogs will not develop PLL as a result of the
mutation we are testing for, although we cannot exclude the
possibility they might develop PLL due to other causes, such as
trauma or the effects of other, unidentified mutations.
• GENETICALLY AFFECTED: these dogs have two copies of the
mutation and will almost certainly develop PLL during their lifetime.
We advise that all genetically affected dogs have their eyes
examined by a veterinary ophthalmologist every 6 months, from the
age of 18 months, so the clinical signs of PLL are detected as early
as possible.
Carriers
•
CARRIER: these dogs have one copy of the mutation and one normal copy
of DNA. Our research has demonstrated that carriers have a very low risk of
developing PLL. The majority of carriers do not develop PLL during their
lives but a small percentage do. We currently estimate that between 2% –
20% of carriers will develop the condition, although we believe the true
percentage is nearer to 2% than 20%. We do not currently know why some
carriers develop the condition whereas the majority do not, and we advise
that all carriers have their eyes examined by a veterinary ophthalmologist
every 6- 12 months, from the age of 2, throughout their entire lives.
Current best estimate = 2.5%
*
Mutation Frequency
• We collected DNA from ‘random’ MBTs to estimate the mutation
frequency
• Samples from 133 dogs collected
• We removed all full & half-siblings – left with 62 dogs that didn’t
share a sire or a dam
– Clear
31 (50%)
– Carrier
29 (47%)
– Affected 2 (3%)
MBTs Tested
Tests Requested
Tests Completed
Clear
%
Carrier
%
Affected
%
Australia
27
2
0
0%
2
100%
0
0%
Czech Republic
3
3
1
33%
2
67%
0
0%
Eire
1
0
Estonia
1
1
0
0%
1
100%
0
0%
Finland
5
3
1
33%
2
67%
0%
France
4
2
0%
2
100%
0%
Germany
57
30
11
37%
17
57%
Hungary
9
5
1
20%
4
80%
0%
Martinique
1
1
0%
1
100%
0%
Spain
6
3
0%
2
67%
1
33%
Sweden
2
0
UK
68
43
27
63%
15
35%
1
2%
Ukraine
3
0
187
93
41
44%
48
52%
4
4%
2
7%
Advice
•
Normally for recessive conditions we advise breeders to mate their carriers to
tested, clear dogs.
•
Puppies don’t have to be DNA tested unless they will be bred with.
•
NOT all the same advice for PLL.
•
Owners DO need to breed with their carriers, because the mutation frequency is so
high.
•
But all puppies that are the offspring of at least one carrier parent should be DNA
tested.
•
*
Dogs carrying the mutation need to be clinically monitored throughout their lives.
Free DNA Test Results
• AHT offers a free DNA test result to owners of all dogs that participated
in the research.
• Dogs that were used were:
– clinically affected dogs
– unaffected dogs over the age of 5yo, with clear eye examination
reports
– Subset of random dogs
197 dogs used in research
41 affected dogs
23 unaffected dogs >5yo
133 ‘random’ dogs
546 free DNA test results have been given out
*
We have been unable to deliver a free DNA test result for 23 dogs
Vitreal Leakage
• As zonules rupture vitreous can leak into the anterior chamber of
the eye – vitreal leakage.
• In some countries evidence of vitreal leak age is sufficient for a dog
to fail an eye examination and be classed as PLL affected
• We have examined samples from several dogs now, including a
small number of MBTs, that have clinical evidence of vitreal leakage
but that are clear of the PLL mutation.
• Vitreal leakage alone is not diagnostic of PLL, although many dogs
with vitreal leakage will progress to develop LL.
Thank You…
• … to all the owners who have provided DNA from their dogs and
supplied us with information.
• … to everybody who has made a financial donation to the research.
We couldn’t have done it without you.